Regional polysterism in the GTP-bound form of the human c-Ha-Ras protein

The backbone 1H, 13C, and 15N resonances of the c-Ha-Ras protein [a truncated version consisting of residues 1−171, Ras(1−171)] bound with GMPPNP (a slowly hydrolyzable analogue of GTP) were assigned and compared with those of the GDP-bound Ras(1−171). The backbone amide resonances of amino acid residues 10−13, 21, 31−39, 57−64, and 71 of Ras(1−171)·GMPPNP, but not those of Ras(1−171)·GDP, were extremely broadened, whereas other residues of Ras(1−171)·GMPPNP exhibited amide resonances nearly as sharp as those of Ras(1−171)·GDP. The residues exhibiting the extreme broadening, except for residues 21 and 71, are localized in three functional loop regions [loops L1, L2 (switch I), and L4 (switch II)], which are involved in hydrolysis of GTP and interactions with other proteins. From the temperature and magnetic field strength dependencies of the backbone amide resonance intensities, the extreme broadening was ascribed to the exchange at an intermediate rate on the NMR time scale. It was shown that the Ras(1−1...